Communication between nerve cells takes place by the release of neurotransmitters at synapses. Signaling can be extremely rapid - transmitting information via hundreds of action potentials per second. It is not possible to synthesize new synaptic vesicles at this pace, so rapid recycling is essential. Although endocytosis at the synapse is at the core of synaptic transmission, the exact mechanism of this process has continued for over 35 years without resolution. Aim 1. In non-neuronal cells, clathrin is the central player in endocytosis. We will test the role of clathrin in synaptic recycling. Aim 2. Adaptins are thought to collect cargo molecules for recycling and connect them to the clathrin coat. We will test the role of these complexes in localizing specific synaptic cargoes and in promoting endocytosis. Aim 3. Synaptotagmin is not only the main calcium sensor in exocytosis, but also a central player in endocytosis at the synapse. We will examine the role of synaptotagmin in recruiting cargo and adapters into the endocytosis pathway. Aim 4. Dynamin is thought to be required for pinching off endocytosing vesicles. However, dynamin is not required in yeast cells, and recent data indicate that dynamin might only be essential during high frequency stimulation at the synapse. We will examine the role of dynamin in synaptic recycling using null and temperature-sensitive dynamin mutations. There is growing evidence that understanding endocytosis will have direct impact on applied health research, since defects in endocytosis may play causative roles in many neuronal diseases, including Huntington's Disease, the ataxias, Parkinson's Disease, Hermansky-Pudlack syndrome, and Alzheimer's Disease. It is our hope that understanding the process of endocytosis may lead to drug therapies for these diseases in the future. Finally, in the course of resolving this biological question, we are developing innovative new techniques that will aid many other researchers to bring new scientific weapons to these and other problems. PUBLIC HEALTH RELEVANCE: Nerve cells communicate via synaptic transmission, but defects in this process can lead to diseases of the nervous system, such as Parkinson's Disease and Alzheimer's Disease. By studying the basic processes of neurotransmission, we hope to understand the causes of neurodegenerative diseases as well as develop novel therapies.